We are analyzing the interactions between virus glycoproteins and their receptors for several enveloped viruses important for human health. Our goals are to define mechanisms of virus fusion and entry, elucidate mechanisms of virus tropism, identify cellular receptors, develop strategies to elicit and detect neutralizing antibodies, and develop novel therapeutic and preventative approaches. 1) Human herpesvirus 8 (HHV-8) is associated with Kaposis sarcoma, a major opportunistic disease associated with HIV infection. We established a specific fusion assay between cells expressing HHV-8 glycoproteins (HHV-8 chronically infected cells, or cells expressing recombinant gB, gH and gL) and target cells presumably expressing the functional receptor. We identified cells that are fusion-permissive (receptor positive) or fusion-nonpermissive (receptor negative). With this information, we devised a functional selection appoach to isolate the cDNA encoding the receptor from a cDNA library derived from fusion-permissive cells. From a human melanoma cell line library, we identified a candidate fusion receptor protein (designated Mel-10), based on its ability to render non-permissive cells permissive for HHV-8 glycoprotein-mediated fusion. Mel-10 m RNA is present at high levels in fusion-permissive cells, and at low or non-detectable levels in nonpermissive cells. Preliminary studies indicate that antibodies raised against peptides derived from putative extracellullar regions of Mel-10 block HHV-8 fusion.These results suggest that we have identified the first fusion receptor for HHV-8. Mel-10 is different from the putative HHV-8 receptors identified by other laboratories based on binding assays. 2) Flaviviruses. We are particularly interested in developing systems to study HCV entry, for use in assaying neutralizing antibodies as well as for isolation of the HCV receptor. To this end, we seek to produce flavivirus subviral particles particles (SVPs) containing the E1 and E2 glycoproteins on their surface. One approach is to exploit a recombinant baculovirus system, which to date is the only approach that has yielded large amounts of purified HCV SVPs. We seek to encapsidate an RNA encoding a reporter gene that would be activated upon entry of the particles into target cells. Because of the unavailability of a cell system that supports HCV replication, we are also analyzing West Nile virus which is much more experimentally acceptable. Methods developed for WNV will be applied to HCV.